Zou Ping, Wu Shu-Yu, Koteiche Hanane A, Mishra Sanjay, Levic Daniel S, Knapik Ela, Chen Wenbiao, Mchaourab Hassane S
Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.
Department of Medicine, Division of Genetic Medicine, Vanderbilt University Medical Center, Nashville, TN, 37232, USA.
Exp Eye Res. 2015 Sep;138:104-13. doi: 10.1016/j.exer.2015.07.001. Epub 2015 Jul 4.
αA- and αB-crystallins are small heat shock proteins that bind thermodynamically destabilized proteins thereby inhibiting their aggregation. Highly expressed in the mammalian lens, the α-crystallins have been postulated to play a critical role in the maintenance of lens optical properties by sequestering age-damaged proteins prone to aggregation as well as through a multitude of roles in lens epithelial cells. Here, we have examined the role of α-crystallins in the development of the vertebrate zebrafish lens. For this purpose, we have carried out morpholino-mediated knockdown of αA-, αBa- and αBb-crystallin and characterized the gross morphology of the lens. We observed lens abnormalities, including increased reflectance intensity, as a consequence of the interference with expression of these proteins. These abnormalities were less frequent in transgenic zebrafish embryos expressing rat αA-crystallin suggesting a specific role of α-crystallins in embryonic lens development. To extend and confirm these findings, we generated an αA-crystallin knockout zebrafish line. A more consistent and severe lens phenotype was evident in maternal/zygotic αA-crystallin mutants compared to those observed by morpholino knockdown. The penetrance of the lens phenotype was reduced by transgenic expression of rat αA-crystallin and its severity was attenuated by maternal αA-crystallin expression. These findings demonstrate that the role of α-crystallins in lens development is conserved from mammals to zebrafish and set the stage for using the embryonic lens as a model system to test mechanistic aspects of α-crystallin chaperone activity and to develop strategies to fine-tune protein-protein interactions in aging and cataracts.
αA-晶状体蛋白和αB-晶状体蛋白是小分子热休克蛋白，它们通过热力学方式结合不稳定的蛋白质，从而抑制其聚集。α-晶状体蛋白在哺乳动物晶状体中高度表达，据推测，它通过隔离易于聚集的老化损伤蛋白以及在晶状体上皮细胞中发挥多种作用，在维持晶状体光学特性方面起着关键作用。在此，我们研究了α-晶状体蛋白在脊椎动物斑马鱼晶状体发育中的作用。为此，我们进行了吗啉代介导的αA-、αBa-和αBb-晶状体蛋白敲低，并对晶状体的总体形态进行了表征。由于这些蛋白质的表达受到干扰，我们观察到晶状体出现异常，包括反射强度增加。在表达大鼠αA-晶状体蛋白的转基因斑马鱼胚胎中，这些异常情况较少见，这表明α-晶状体蛋白在胚胎晶状体发育中具有特定作用。为了扩展和证实这些发现，我们构建了一个αA-晶状体蛋白敲除斑马鱼品系。与通过吗啉代敲低观察到的情况相比，母本/合子αA-晶状体蛋白突变体中晶状体表型更加一致且严重。大鼠αA-晶状体蛋白的转基因表达降低了晶状体表型的外显率，而母本αA-晶状体蛋白的表达减轻了其严重程度。这些发现表明，α-晶状体蛋白在晶状体发育中的作用从哺乳动物到斑马鱼是保守的，为利用胚胎晶状体作为模型系统来测试α-晶状体蛋白伴侣活性的机制方面以及制定微调衰老和白内障中蛋白质-蛋白质相互作用的策略奠定了基础。
